Protein resonance assignment by solid-state NMR based on H-1-detected C-13 double-quantum spectroscopy at fast MAS

Solid-state NMR spectroscopy is a powerful technique to study insoluble and non-crystalline proteins and protein complexes at atomic resolution. The development of proton (H-1) detection at fast magic-angle spinning (MAS) has considerably increased the analytical capabilities of the technique, enabling the acquisition of H-1-detected fingerprint experiments in few hours. Here an approach based on double-quantum (DQ) C-13 spectroscopy, detected on H-1, is proposed for fast MAS regime (> 60 kHz) to perform the sequential assignment of insoluble proteins of small size, without any specific deuteration requirement. By combining two three-dimensional H-1 detected experiments correlating a C-13 DQ dimension respectively to its intra-residue and sequential (15) N-H-1 pairs, a sequential walk through DQ (Ca + CO) resonance is obtained. The approach takes advantage of fast MAS to achieve an efficient sensitivity and the addition of a DQ dimension provides spectral features useful for the resonance assignment process.

Automated summary

The approach combines double-quantum C-13 spectroscopy and fast MAS to achieve sequential assignment of insoluble proteins, providing efficient sensitivity and useful spectral features for resonance assignment process.